Project Summary Proteins from the Fanconi anemia (FA) pathway play an integral role in DNA repair by homologous recombination (HR). FA is a multigenic disorder marked by progressive bone marrow failure and a strong cancer predisposition. Numerous studies have linked mutations in FA genes to familial breast, pancreatic, and other cancers, and have also provided ample evidence to implicate silencing of FA genes in the etiology of sporadic cancers. FA cells are hypersensitive to radiation and other DNA damaging agents, DNA crosslinking chemicals and reactive aldehydes in particular, prone to DNA replicative stress, and exhibit chromosome fragility. These phenotypic manifestations stem from defects in DNA damage signaling and repair, and FA protein functional and physical interactions have indicated an important link to the familial breast cancer proteins BRCA1 and BRCA2. The involvement of the FA/BRCA-dependent DNA damage response in cancer suppression underscores the need to understand the mechanistic underpinnings of this genome maintenance pathway. In this project, we will employ a combination of biochemical and in vivo approaches to test the novel hypotheses that the FA pathway coordinates the prevention and resolution of genotoxic structures resulting from aberrant transcription events and HR. Aim 1 will focus on the mechanism whereby the ID2 complex and UAF1-containing complexes engage nucleic acid containing structures and activate HR. Aim 2 will explore how the FANCM-BLM axis resolves pathogenic RNA containing and other nucleic acid intermediates in a way that regulates HR activities. The success of this project is assured by the complementary expertise of the two participating Yale groups, led by Dr Patrick Sung and Dr Gary Kupfer, and an exceptionally strong collaborative framework within the broader Yale community. In addition, the PIs have enlisted two investigators, Dr. Claudia Wiese and Dr. Andres Aguilera, whose expertise will even more elegantly allow the team to interrogate this novel area of genomic instability. These attributes help ensure that findings of the highest possible impact will be obtained. Since the biology of FA intersects with cancer biology in general, our project promises to shed light on critical processes of genomic surveillance as well as common themes of oncogenesis. We expect our studies to yield insight into common pathways of cancer and to identify novel targets for manipulation in cancer therapy.